The principal goal of this research is to study the enzymology of replication of yeast chromosomal DNA. The work will involve two aspects in DNA replication. In addition to studying replication systems in cell extracts that are specific for yeast replication origins, special attention will be paid to the purification and exact characterization of the enzymes involved in the elongation stage of replication: i.e. the DNA polymerases, the primase and the single-stranded DNA binding protein(s). Previous reports on the yeast DNA polymerases have to be reexamined in view of the fact that these studies almost certainly have been performed on proteolytic fragments. In analogy with the E. coli DNA polymerase III holoenzyme, a search will be made for a yeast holoenzyme using specific assay systems. The yeast primase willbe purified and we will determine whether there are conditions which make the priming event specific for particular single-stranded yeast DNA sequences. These purified replication proteins will be used in assay systems that measure replication from yeast replication origins. Replication mutants will be studied in vitro in these several assay systems. The proposed study of eucaryotic replication will focus on yeast because, of all eucaryotic organisms, yeast is the one most amenable to biochemical and genetic manipulation. An intensive study of DNA replication and its regulation in yeast will undoubtedly be of significance in understanding cell growth and division in higher organisms.